Antiproliferative agents

ABSTRACT

This invention relates to certain dioxodihydropyridine derivatives and certain methacrylic acid and furan derivative precursors thereof, processes for their preparation, pharmaceutical compositions containing such compounds and their use as anti-proliferative agents, especially tumour growth inhibitors and anti-cancer agents, antibiotics and/or antiviral agents.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of PCT GB02/01119, filed Dec. 3,2002, and published in English on Sep. 19, 2002 as WO 02/072553. PCTGB02/01119 claimed the priority of British application 0106137.3, filedDec. 3, 2001. The disclosures of both are incorporated herein byreference.

FIELD OF THE INVENTION

This invention relates to certain dioxodihydropyridine derivatives andcertain methacrylic acid and furan -derivative precursors thereof,processes for their preparation, pharmaceutical compositions containingsuch compounds and their use as anti-proliferative agents, especiallytumour growth inhibitors and anti-cancer agents, antibiotics and/orantiviral agents.

BACKGROUND OF THE INVENTION

Various anti-cancer agents are known which are currently in clinicaluse. Some of these, such as cyclophosphamide and chlorambucil, arederived from the highly toxic group of N-Lost derivatives which containa so-called N-Lost group, that is, a bis(2-chloroethyl)amino group. Theoriginal N-Lost derivatives are known to have cancer-triggeringproperties and were formerly used as chemical weapons. However,clinically acceptable compounds, such as cyclophosphamide andchlorambucil, have been prepared by modifying the original N-Loststructure to produce analogues with a significantly reduced toxicity.Nevertheless, these analogues all contain chemically reactive chlorinewhich is believed to be essential for their biological activity. Incontrast, N-methylformamide is known as an anti-cancer agent butcontains no chemically reactive chlorine.

SUMMARY AND DETAILED DESCRIPTION OF THE INVENTION

The present invention utilises the N-methylformamide moiety, sometimesin conjunction with the feature of chemically reactive halogen,especially chlorine, to produce biologically active compounds which arestructurally distinct from the compounds discussed above. According tothe present invention there is therefore provided a compound of thegeneral formula

in which R¹ represents a hydrogen atom or an optionally substitutedalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, aralkyl,heterocyclyl or heterocyclylalkyl group; R² represents a hydrogen atomor an optionally substituted alkyl, alkenyl, alkynyl, cycloalkyl,cycloalkenyl, aryl, aralkyl, acyl, heterocyclyl or heterocyclylalkylgroup; and W and X each independently represents a hydrogen or halogenatom.

Any alkyl, alkenyl or alkynyl group, unless otherwise specified, may belinear or branched and may contain up to 12, preferably up to 6, andespecially up to 4 carbon atoms. Preferred alkyl groups are methyl,ethyl, propyl and butyl. Preferred alkenyl and alkynyl groups includepropenyl, butenyl, propynyl and butynyl groups. When an alkyl moietyforms part of another group, for example the alkyl moiety of an aralkylgroup, it is preferred that it contains up to 6, especially up to 4,carbon atoms. Preferred alkyl moieties are methyl and ethyl.

An aryl group may be any aromatic hydrocarbon group and may contain from6 to 24, preferably 6 to 18, more preferably 6 to 16, particularly 6 to14, and especially 6 to 10 carbon atoms. Preferred aryl groups includephenyl, naphthyl, anthryl, phenanthryl and pyryl groups, especially aphenyl or naphthyl, and particularly a phenyl, group. When an arylmoiety forms part of another group, for example the aryl moiety of anaralkyl group, it is preferred that it is a phenyl, naphthyl, anthryl,phenanthryl or pyryl, especially phenyl or naphthyl, and particularly aphenyl, moiety.

An aralkyl group may be any alkyl group substituted by an aryl group. Apreferred aralkyl group contains from 7 to 30, particularly 7 to 24 andespecially 7 to 18, carbon atoms, particularly preferred aralkyl groupsbeing benzyl, naphthylmethyl, anthrylmethyl, phenanthrylmethyl andpyrylmethyl groups. A particularly preferred aralkyl group is a benzylgroup.

A cycloalkyl group may be any saturated cyclic hydrocarbon group and maycontain from 3 to 12, preferably 3 to 6, and especially 3 to 6, carbonatoms. Preferred cycloalkyl groups are cyclopropyl, cyclopentyl andcyclohexyl groups.

A cycloalkenyl group may be any cyclic hydrocarbon group which containsat least one carbon-carbon double bond. Thus, a cycloalkenyl group iseffectively a cycloalkyl group in which at least one carbon-carbonsingle bond has been replaced by a carbon-carbon double bond. Acycloalkenyl group may therefore contain from 3 to 12, preferably 3 to8, and especially 3 to 6, carbon atoms. Preferred cycloalkenyl groupsare cyclopentenyl-and cyclohexenyl groups.

An acyl group may be any group of the general formula R—CO— where Rrepresents an optionally substituted alkyl, alkenyl, alkynyl,cycloalkyl, cycloalkenyl, aryl, aralkyl, heterocyclyl orheterocyclylalkyl group. Preferred acyl groups are alkanoyl, especiallyC₁₋₄ alkanoyl, and aroyl, especially benzoyl, groups.

A heteroaryl group may be any aromatic monocyclic or polycyclic ringsystem which contains at least one heteroatom. Preferably, a heteroarylgroup is a 5- to 18-membered, particularly a 5- to 14-membered, andespecially a 5- to 10-membered, aromatic ring system containing at leastone heteroatom selected from oxygen, sulphur and nitrogen atoms.Preferred heteroaryl groups include pyridyl, pyrylium, thiopyrylium,pyrrolyl, furyl, thienyl, indolinyl, isoindolinyl, indolizinyl,imidazolyl, pyridonyl, pyronyl, pyrimidinyl, pyrazinyl, oxazolyl,thiazolyl, purinyl, quinolinyl, isoquinolinyl, quinoxalinyl,pyridazinyl, benzofuranyl, benzoxazolyl and acridinyl groups.

A heterocyclic group may be any monocyclic or polycyclic ring systemwhich contains at least one heteroatom and may be unsaturated orpartially or fully saturated. The term “heterocyclic” thus includesheteroaryl groups as defined above as well as non-aromatic heterocyclicgroups. Preferably, a heterocyclic group is a 3- to 18-membered,particularly a 3- to 14-membered, especially a 5- to 10-membered, ringsystem containing at least one heteroatom selected from oxygen, sulphurand nitrogen atoms. Preferred heterocyclic groups include the specificheteroaryl groups named above as well as pyranyl, piperidinyl,pyrrolidinyl, dioxanyl, piperazinyl, morpholinyl, thiomorpholinyl,morpholinosulphonyl, tetrahydroisoquinolinyl and tetrahydrofuranylgroups.

A heterocyclylalkyl group may be any alkyl group substituted by aheterocyclic group. Preferably, the heterocyclic moiety is a 3- to 18-membered, particularly a 3- to 14-membered, and especially a 5- to10-membered, heterocyclic group as defined above and the alkyl moiety isa C₁₋₆ alkyl, preferably C₁₋₄ alkyl, and especially methyl, group.

When any of the foregoing substituents are designated as beingoptionally substituted, the substituent groups which are optionallypresent may be any one or more of those customarily employed in thedevelopment of pharmaceutical compounds and/or the modification of suchcompounds to influence their structure/activity, stability,bioavailability or other property. Specific examples of suchsubstituents include, for example, halogen atoms, nitro, cyano,hydroxyl, cycloalkyl, alkyl, alkenyl, haloalkyl, alkoxy, haloalkoxy,amino, alkylamino, dialkylamino, formyl, alkoxycarbonyl, carboxyl,alkanoyl, alkylthio, alkylsulphinyl, alkylsulphonyl, alkylsulphonato,arylsulphinyl, arylsulphonyl, arylsulphonato, carbamoyl, alkylamido,aryl and aralkyl groups.

When any of the foregoing optional substituents represents or containsan alkyl or alkenyl substituent group, this may be linear or branchedand may contain up to 12, preferably up to 6, and especially up to 4,carbon atoms. A cycloalkyl group may contain from 3 to 8, preferablyfrom 3 to 6, carbon atoms. An aryl group or moiety may contain from 6 to10 carbon atoms, phenyl groups being especially preferred. A halogenatom may be a fluorine, chlorine, bromine or iodine atom and any groupwhich contains a halo moiety, such as a haloalkyl group, may thuscontain any one or more of these halogen atoms.

Preferred optional substituents include halogen atoms, nitro, cyano,hydroxyl, C₁₋₆ alkyl, C₁₋₆ haloalkyl, C₁₋₆ alkoxy, C₁₋₆ haloalkoxy,amino, C₁₋₆ alkylamino, di-(C₁₋₆ alkyl)amino, formyl, C₁₋₆alkoxycarbonyl, carboxyl, C₁₋₆ alkanoyl, C₁₋₆ alkylthio, C₁₋₆alkylsulphinyl, C₁₋₆ alkylsulphonyl, carbamoyl and C₁₋₆ alkylamidogroups. Particularly preferred optional substituents include halogenatoms, nitro, cyano,:hydroxyl, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxyand C₁₋₄ haloalkoxy groups with halogen atoms being especiallypreferred.

Preferably, R¹ represents a C₁₋₁₂ alkyl or C₆₋₁₄ aryl group, each groupbeing optionally substituted by one or more substituents selected fromthe group consisting of halogen atoms, nitro, cyano, hydroxyl, C₁₋₄alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxy and C₁₋₄ haloalkoxy groups. Morepreferably, R¹ represents a C₁₋₆ alkyl, especially a C₁₋₄ alkyl, groupoptionally substituted by one or more substituents selected from thegroup consisting of halogen atoms, nitro, cyano, hydroxyl, C₁₋₄ alkoxyand C₁₋₄ haloalkoxy groups. It is particularly preferred that R¹represents an unsubstituted C₁₋₄ alkyl group, especially a methyl group.

It is preferred that R² represents a hydrogen atom or a C₁₋₁₂ alkyl orC₆₋₁₄ aryl group, each group being optionally substituted by one or moresubstituents selected from the group consisting of halogen atoms, nitro,cyano, hydroxyl, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxy and C₁₋₄haloalkoxy groups. More preferably, R² represents a hydrogen atom or aC₁₋₆ alkyl, especially a C₁₋₄ alkyl, group optionally substituted by oneor more substituents selected from the group consisting of halogenatoms, nitro, cyano, hydroxyl, C₁₋₄ alkoxy and C₁₋₄ haloalkoxy groups.It is particularly preferred that R² represents a hydrogen atom or anunsubstituted C₁₋₄ alkyl, especially a methyl group. Most preferably, R²represents a hydrogen atom.

Preferably, W and X each independently represents a hydrogen, chlorineor bromine atom, especially a hydrogen or chlorine atom. Morepreferably, W and X are both the same.

In a particularly, preferred sub-group of compounds of formula I, R¹represents a methyl group, R² represents a hydrogen atom and W and Xboth represent a hydrogen atom or both represent a chlorine atom.

It should be appreciated that compounds of general formula I in which R²represents a hydrogen atom can tautomerise. The three main tautomericforms are set out below:

The present invention thus includes both the individual tautomers andmixtures of such tautomers.

Particularly preferred compounds include3,4-dichloro-6-hydroxy-1-methyl-2,5-dioxo-1,6-dihydropyridine (alsoknown as 3,4-dichloro-6-hydroxy-1-methyl-1,6-dihydropyridine-2,5-dione)and 6-hydroxy-1-methyl-2,5-dioxo-1,6-dihydropyridine (also known as6-hydroxy-1-methyl-1,6-dihydropyridine-2,5-dione), with the formercompound being especially preferred.

The invention also provides a process for the preparation of a compoundof the general formula I as defined above which comprises reacting acompound of the general formula

in which R¹, R², W and X are as defined above and Y represents ahydrogen atom, a hydroxyl group or a group —OM where M represents analkali metal atom, with a base in the presence of a compound of thegeneral formula

in which R¹ and R² are as defined above.

It is preferred that Y represents a hydrogen atom, a hydroxyl group or agroup —OM where M represents a sodium or potassium atom. Morepreferably, Y represents a hydroxyl group.

The base may be an organic or inorganic base. Suitable organic basesinclude pyridine and tertiary amines, such as triethylamine. Suitableinorganic bases include carbonates and hydrogencarbonates of alkalimetals and alkaline earth metals, such as sodium, potassium, lithium,calcium, magnesium, etc. In general, inorganic bases are preferred withsodium hydrogencarbonate being especially preferred.

Preferably, the reaction is carried out in the presence of an acylhalide. Suitable acyl halides include C₂₋₄ acyl halides and halo C₂₋₄acyl halides, with acetyl chloride and dichloroacetyl chloride beingespecially preferred.

The reaction is preferably carried out at a temperature from 0 to 80°C., with 60° C. being particularly preferred.

A possible reaction scheme for this process is set out in Scheme Ibelow:

On the basis of the above proposed reaction scheme, it may be necessaryor desirable to convert any compound of formula IIB to the correspondingcompound of formula IIA to facilitate the reaction. This can be readilyachieved by exposure to ultra violet light. Thus, exposure to ultraviolet light is an optional additional step in the process of theinvention.

The invention also provides a process for the preparation of a compoundof the general formula IIA or IIB as defined above which comprisesreacting a compound of the general formula

in which R¹, R², W and X are as defined above and R³ represents anoptionally substituted alkyl, aryl or aralkyl group, with a base.

This process produces a mixture of compounds of formula IIA and IIB.Under normal conditions, this process will produce a mixture containingabout 80% compounds of formula IIA and about 20% compounds of formulaIIB. However, the quantity of compounds of formula IIB can be increasedby using more heat in the process and the quantity of compounds offormula IIA can be increased by exposure to ultra violet light.

Preferably, R³ represents a C₁₋₆ alkyl, especially a C₁₋₄ alkyl, groupoptionally substituted by one or more substituents selected from halogenatoms, nitro, cyano, hydroxyl, C₁₋₄ alkoxy and C₁₋₄ haloalkoxy groups,with halogen atoms, especially chlorine atoms, being particularlypreferred as optional substituents. More preferably, R³ represents aC₁₋₄ alkyl group optionally substituted by one or more halogen,especially chlorine, atoms. It is especially preferred that R³represents a chloromethyl or dichloromethyl group

The base may be an organic or inorganic base. Suitable organic basesinclude pyridine and tertiary amines, such as triethylamine. Suitableinorganic bases include carbonates and hydrogencarbonates of alkalimetals and alkaline earth metals, such as sodium, potassium, lithium,calcium, magnesium, etc. In general, inorganic bases are preferred withsodium hydrogencarbonate being especially preferred.

The reaction is preferably carried out at a temperature of 0 to 80° C.,with 60° C. being particularly preferred.

According to another aspect of the invention, compounds of formula IV asdefined above can be prepared by reacting a compound of the generalformula

in which R¹ and R² are as defined above, with a compound of the generalformula

in which Z¹ and Z² each independently represents a hydrogen or halogen,especially a chlorine, atom and Z³ is a halogen, preferably a chlorine,atom.

Preferably, the compound of general formula IIII is reacted with anexcess of the compound of general formula V. More preferably, the molarratio of the compound of general formula V to the compound of generalformula IIII is at least 2:1. Thus, at least two equivalents of thecompound of general formula V are used for each equivalent of thecompounds of the general formula III.

Preferably, the reaction is carried out at a temperature of from 0° C.to the reflux temperature of the reaction mixture, especially at 60° C.to the reflux temperature of the reaction mixture. In general, anincrease in reaction temperature results in an increase in the yield ofthe desired product of general formula IV.

Compounds of the general formula IIII and V are known compounds or canbe prepared from known compounds by processes analogous to knownprocesses.

A reaction scheme for this process is set out in Scheme II below:

This process provides a general method for synthesising 2,5-substitutedfuran derivatives in a one-pot reaction. However, if desired, the amideintermediate of the general formula V can be isolated.

The processes set out above describe the preparation of compounds of thegeneral formulae I, IIA, IIB and IV in separate steps. However,compounds of formula I can be prepared in a one-pot reaction startingfrom compounds of formulae III and V and this may be particularlyadvantageous.

The compounds of general formula IIA and IIB are novel. According toanother aspect of the invention there is therefore provided a compoundof the general formula

or a salt thereof, in which R¹, R², W, X and Y are as defined above.

Suitable salts include acid addition salts and these may be formed byreaction of a suitable compound of formula IIA or IIB with a suitableacid, such as an organic acid or a mineral acid. Acid addition saltsformed by reaction with a mineral acid are particularly preferred,especially salts formed by reaction with hydrochloric or hydrobromicacid.

It should be appreciated that the compounds of general formula IIA andIIB are geometric isomers of each other. The present invention thusincludes both the individual isomers and mixtures of such isomers.

Particularly preferred compounds include (E)- and(Z)-2,3-dichloro-4-[N-formyl-N-methylamino]-4-oxo-but-2-enoic acid (alsoknown as (E)- and(Z)-4-(N-methylformamido)-4-oxo-2,3-dichloromethacrylic acid) and (E)-and (Z)-4-[N-formyl-N-methylamino]-4-oxo-but-2-enoic acid (also known as(E)- and (Z)-4-(N-methylformamido)-4-oxo-methacrylic acid), with theformer compounds being especially preferred. The Z-isomers of thesecompounds are particularly preferred.

The compounds of general formula IV are also novel and the inventiontherefore further provides a compound of the general formula

in which R¹, R², R³, W and X are as defined above. Preferred compoundsof formula IV include5-[N-formyl-N-methylamino]-3,4-dichloro-2-furyl-2,2-dichloroacetate(also known as3,4-dichloro-2-(N-methylformamido)-5-dichloroacetoxyfuran) and5-[N-formyl-N-methylamino]-2-furyl-2-chloroacetate (also known as2-(N-methylformamido)-5-chloroacetoxyfuran), with the former compoundbeing especially preferred.

The compounds of general formulae I, IIA and IIB have been found to beeffective in the treatment and/or prophylaxis of a variety of diseases,particularly disorders involving cell proliferation, such as cancer,cancer metastasis and the growth of non-malignant tumours, as well asdiseases caused by bacteria or other microbes and viral diseases. Thesecompounds exhibit a wide spectrum of anti-cancer activity. However, theyare particularly suitable for the treatment of lung, ovary and/or coloncancer. Moreover, since the compounds of general formula IV can bereadily converted into compounds of general formula IIA or IIB in thepresence of a base, the compounds of general formula IV function asprodrugs. The term “prodrug” refers to a compound whose metabolitepossesses therapeutic utility greater than the compound itself. Themetabolite arises from chemical and/or enzymatic transformations in vivofollowing administration of the prodrug.

In view of the above, the present invention also provides a compound ofthe general formula I as defined above and/or a compound of the generalformula IIA or IIB or a salt thereof as defined above and/or a compoundof the general formula IV as defined above for use in medicine,particularly as an anti-proliferative agent, an antibiotic or ananti-viral agent, and especially as a tumour growth inhibitor and/oranti-cancer agent.

The invention also includes the use of a compound of the general formulaI as defined above and/or a compound of the general formula IIA or IIBor a salt thereof as defined above and/or a compound of the generalformula IV as defined above for the manufacture of a medicament for useas an anti-proliferative agent, an antibiotic or an anti-viral agent,and especially as a tumour growth inhibitor and/or anti-cancer agent.

In another aspect, the invention provides a method for treating orpreventing a disorder involving cell proliferation, a disease caused bya bacterium or a microbe or a viral disease which comprisesadministering to a patient a therapeutically or prophylacticallyeffective amount of a compound of the general formula I as defined aboveand/or a compound of the general formula IIA or IIB or a salt thereof asdefined above and/or a compound of the general formula IV as definedabove. Preferably, the disorder involving cell proliferation is cancer,cancer metastasis or the growth of a non-malignant tumour.

In a further aspect, the invention provides a compound of the generalformula IV as defined above for use as a prodrug. The use of a compoundof the general formula IV as defined above for the manufacture of amedicament for use as a prodrug is also provided.

The invention also provides a pharmaceutical composition which comprisesa carrier and, as active ingredient, a compound of the general formula Ias defined above and/or a compound of the general formula IIA or IIB ora salt thereof as defined above and/or a compound of the general formulaIV as defined above. A process for the preparation of a pharmaceuticalcomposition as defined above is also provided which comprises bringing acompound of the general formula I as defined above and/or a compound ofthe general formula IIA or IIB or a salt thereof as defined above and/ora compound of the general formula IV as defined above into associationwith a carrier.

A pharmaceutically acceptable carrier may be any material with which theactive ingredient is formulated to facilitate administration. A carriermay be a solid or a liquid, including a material which is normallygaseous but which has been compressed to form a liquid, and any of thecarriers normally used in formulating pharmaceutical compositions may beused. Preferably, compositions according to the invention contain 0.5 to95% by weight of active ingredient.

The compounds of general formula I, general formula IIA and IIB andgeneral formula IV can be formulated as, for example, tablets, capsules,suppositories or solutions. These formulations can be produced by knownmethods using conventional solid carriers such as, for example, lactose,starch or talcum or liquid carriers such as, for example, water, fattyoils or liquid paraffins. Other carriers which may be used includematerials derived from animal or vegetable proteins, such as thegelatins, dextrins and soy, wheat and psyllium seed proteins; gums suchas acacia, guar, agar, and xanthan; polysaccharides; alginates;carboxymethylcelluloses; carrageenans; dextrans; pectins; syntheticpolymers such as polyvinylpyrrolidone; polypeptide/protein orpolysaccharide complexes such as gelatin-acacia complexes; sugars suchas mannitol, dextrose, galactose and trehalose; cyclic sugars such ascyclodextrin; inorganic salts such as sodium phosphate, sodium chlorideand aluminium silicates; and amino acids having from 2 to 12 carbonatoms such as a glycine, L-alanine, L-aspartic acid, L-glutamic acid,L-hydroxyproline, L-isoleucine, L-leucine and L-phenylalanine.

Auxiliary components such as tablet disintegrants, solubilisers,preservatives, antioxidants, surfactants, viscosity enhancers, colouringagents, flavouring agents, pH modifiers, sweeteners or taste-maskingagents may also be incorporated into the composition. Suitable colouringagents include red, black and yellow iron oxides and FD & C dyes such asFD & C blue No. 2 and FD & C red No. 40 available from Ellis & Everard.Suitable flavouring agents include mint, raspberry, liquorice, orange,lemon, grapefruit, caramel, vanilla, cherry and grape flavours andcombinations of these. Suitable pH modifiers include sodiumhydrogencarbonate, citric acid, tartaric acid, phosphoric acid,hydrochloric acid and maleic acid. Suitable sweeteners includeaspartame, acesulfame K and thaumatin. Suitable taste-masking agentsinclude sodium hydrogencarbonate, ion-exchange resins, cyclodextrininclusion compounds, adsorbates or microencapsulated actives.

The inclusion of a compound which is capable of generating an alkalineenvironment in the body or the co-administration of a base may beparticularly advantageous when the composition contains a compound offormula IV to encourage the conversion of such a compound into acompound of formula IIA or IIB.

For treatment of and prophylaxis against disorders involving cellproliferation, diseases caused by bacteria or microbes or viraldiseases, amounts of 0.5 to 100, preferably 20 to 50, mg/kg body weightactive compound are preferably administered daily to obtain the desiredresults. Nevertheless, it may be necessary from time to time to departfrom the amounts mentioned above, depending on the body weight of thepatient, the method of application, the animal species of the patientand its individual reaction to the drug or the kind of formulation orthe time or interval in which the drug is applied. In special cases, itmay be sufficient to use less than the minimum amount given above,whilst in other cases the maximum dose may have to be exceeded. For alarger dose, it may be advisable to divide the dose into several smallersingle doses.

The invention is further illustrated by the following examples.

EXAMPLE 1 Preparation of N-methyl-N-formyl-α,α-dichloroacetamide

(Formula VI: R¹═—CH₃; R²═—H; Z=Cl)

25 g of dichloroacetyl chloride (1.25 eq.) was added to a solution of 50g N-methylformamide (NMF) in chloroform at ambient temperature. Thereaction mixture was washed with an aqueous solution of sodiumhydrogencarbonate, dried with magnesium sulfate and the solvent wasevaporated off in vacuum to give N-methyl-N-formyl-α,α-dichloroacetamide(86% yield) in high purity as a yellow oil. The chemical structure wasconfirmed by APCl-MS, FT-IR, 1H and 13C magnetic resonance spectroscopy.

¹H-NMR (DMSO-d₆): 3.11 (d, 3H, NCH₃, J=4.7 Hz); 6.58+5.93 (s, 1H,COCHCl₂); 9.19 (br.s., 1H, NCOH); ¹³C-NMR (DMSO-d₆): 25.36+26.78;64.23+64.83; 162.95, 165.44+166.32; IR (KBr): 3394, 1702, 1667, 1162,670; MS (APCl+, m/z): 171 (M⁺+1).

N-methyl-α,α-dichloroacetamide precipitated out within a couple of daysin a freezer as white needles (48% yield). The chemical structure wasconfirmed by APCl-MS, FT-IR, 1H and 13C magnetic resonance spectroscopy.

¹H-NMR (DMSO-d₆): 2.67 (d, 3H, NCH₃, J=4.7 Hz.); 6.44 (s, 1H, COCHCl₂);8.51 (br.s., 1H, NH); ¹³C-NMR (DMSO-d₆): 26.78, 67.32, 164.53; IR (KBr):3394, 1666, 1162, 670; MS (APCl+, m/z): 143 (M⁺+1).

EXAMPLE 2 Preparation of5-[N-formyl-N-methylamino]-3,4-dichloro-2-furyl-2,2-dichloroacetate(also known as3,4-dichloro-2-(N-methylformamido)-5-dichloroacetoxvfuran) (One-potsynthesis)

(Formula IV: R¹═—CH₃; R²═—H; R³═—CHCl₂; W═X═Cl.)

60 ml of NMF were added in portions to 140 ml of dichloroacetylchloride. After the first addition of NMF, the temperature was increasedand the addition of NMF was continued in portions, so that the mixtureproceeded to reflux gently. The reaction was completed after severalhours. For workup, the reaction mixture was poured onto ice and theremaining oily residue was washed repeatedly with water. The oil wasdissolved in dichloromethane (DCM) and the solution of the furan in DCMwas washed many times with water. The organic phase was dried withmagnesium sulfate and the solvent was carefully removed at roomtemperature in vacuum. The title compound was obtained as pure,yellow-orange oil (72% yield). The chemical structure of the furan wasconfirmed by APCl-MS, FT-IR, 1H and 13C magnetic resonance spectroscopy.

¹H-NMR (CDCl₃-d₆): 9.28 (s, 1H, CHO), 6.52 (s, 1H, ClClH), 3.19 (s, 3H,N-Me);

¹³C-NMR (CDCl₃-d₆): 165.55, 165.19+162.12 (CHO), 165.09, 165.03(COClCl), 157.86, 140.12, 66.10+53.60 (C—ClCl), 28.45+28.07 (N-Me: MS(ACPI+, m/z): 342, 341, 340 [(M⁺+1)+OH], 324, 323, 322 (M+1), 212, 210[(M+1)-OAc], 183, 182 (210-CO)

EXAMPLE 3 Preparation of(Z)-2,3-dichloro-4-[N-formyl-N-methylamino]-4-oxo-but-2-enoic acid (alsoknown as (Z)-4-(N-methylformamido)-4-oxo-2,3-dichloromethacrylic acid)

(Formula IIA: R¹═—CH₃; R²═—H; W═X═Cl; Y═—OH)

2 g of 5-[formyl(methyl)amino]-3,4-dichloro-2,2-dichloroacetate preparedas described in Example 2 were mixed with 1.5 g of solid sodiumhydrogencarbonate and 5 ml of water. The mixture was shaken vigorouslyunder development of carbon dioxide. After 3 hours a homogeneoussolution of the title compound in water was obtained and the sodiumdichloroacetate which precipitated out was filtered off. Water was addedto this solution until a total volume of 10 ml was obtained. Thissolution was then used to evaluate the efficacy of this compound inanimal tumour models (see Example 4 below).

Content: 10%

10 mg in 100 μl

The title compound was independently isolated as a non-distillableviscous, water-soluble liquid. The chemical structure was confirmed byAPCl-MS, FT-IR, 1H and 13C magnetic resonance spectroscopy.

¹H-NMR (D₂O), MeOD-d₆): 7.77 (s, 1H, CHO), 2.53 (s, 3H, Me) ₁₃; C-NMR(D₂O, MeOD-d₆): 169.13, 166.32, 165.14, 163.04, 162.95, 24.33 (Me); MS(APCl+, m/z): 241,239 (OMe), 229, 230 (OH), 210,184 (—CO)

EXAMPLE 4 Preparation of3,4-dichloro-6-hydroxy-1-methyl-2,5-dioxo-1.6-dihydropyridine (alsoknown as 3,4-dichloro-6-hydroxy-1-methyl-1,6-dihydropyridine-2,5-dione)(One-pot synthesis) (Formula I: R¹═CH₃; R²═H; W═X═Cl

60 ml NMF were added in portions to 140 ml of dichloroacetyl chloride.After the first addition of NMF, the temperature was increased and theaddition of NMF was continued in portions, so that the mixture proceededto reflux gently. The reaction was completed within a couple of hours.

Method A

20 g of the reaction mixture was mixed with 15 g solid sodiumhydrogencarbonate and 50 ml of water. The reaction mixture was extractedwith ether in a Ludwig (continuous extraction unit) extractor overnight.The ether solution was dried with magnesium sulphate and was evaporatedoff under reduced pressure. The remaining yellow oil was distilled in afine vacuum to give the title compound in 40% yield.

Method B

15 g of solid sodium hydrogencarbonate was added to 20 g of the crudematerial. A small amount of water was added followed by ethyl acetate assolvent. If the reaction rate decreased, more water was added. After thereaction was completed, the solution was dried with magnesium sulphateand the solvent was distilled off. The distillation of the remaining oilgave the title compound in 35% yield.

¹H-NMR (DMSO-d₆): 7.6 (bs, 1H, OH); 6.0 (s, 1H)

¹³C-NMR (DMSO-d₆): 25.36+26.78; 102.5 (C6); 123.44, 126.6 (C3, C4), 156,162 (CO); IR (KBr): 3610, 3394, 1702, 1715, 1667, 1162, 670;

MS (APCl+, m/z): 210 (M⁺+1)

EXAMPLE 5

Assessment of Tumour Growth Inhibition

Pure strain NMRI mice (age 6-8 weeks) from our inbred colony were usedfor transplanting MAC (murine colon cancer) tumours, M5076 reticulumcell sarcoma (ovarian) were transplanted in BDF1 mice purchased fromHarlan, Bicester, U.K. Animals were fed on RM3E diet (Lillco-England)and water ad libitum.

MAC 13 and M5076

Approximately 2×2 mm of MAC13 tumour fragments were transplantedsubcutaneously in the inguinal region via a trocar while the M5076 washomogenised and 1×10⁶ cells transplanted intramuscularly in the inguinalregion. Tumour bearing mice were randomised in groups of 5-7 animals pergroup. Treatment commenced one day after transplant for M5076 and twodays for MAC13. Untreated or vehicle controls were included. Dailyweights were recorded. Chemotherapy effects were assessed on day 14 orthereabouts depending on the tumour sizes and condition of the animals.Mice were killed and the effects of the chemotherapy were assessed bythe differences in tumour weights and expressed as:%T/C, i.e. inhibition=100-Treated weight/Control weight×100%Body weight changes were recorded in order to assess toxicity.MAC15A

The tumour is routinely passaged as an ascites. Approximately 2×10⁵cells were transplanted subcutaneously in saline in a volume of 0.2 ml.Mice were randomised in groups of 5-7 animals per group. Untreated orvehicle controls were included. Treatment commenced two days aftertransplant. Body weight was recorded daily. The effects of chemotherapywere assessed between 7-9 days after transplant. Mice were killed andthe effects measured by the differences in tumour weights and expressedas:%T/C, i.e. inhibition=Treated weight/Control weight×100%Body weight changes were recorded in order to assess toxicity.

The results are set out in Tables 1, 2, 3 and 4 below TABLE 1 The effectof the compound of Example 3 on the growth of MAC 13 tumour, murinecolon cancer, transplanted s.c. on NMRI mice and treatment given byintraperitoneal route Dose: % mg/kg Tumour Tumour Tumour Tumour TumourTumour Mean Tumour body weight weight weight weight weight weight tumourgrowth Days of weight (g) (g) (g) (g) (g) (g) weight sem inhibitiontreatment 400 0.053 0.045 0.030 0.023 0.0 0.036 0.031 0.007 92 1-5Control 0.413 0.304 0.270 0.538 0.641 0.209 0.395 0.068 0 — untreated

TABLE 2 The effect of the compound of Example 3 on the growth of M5076tumour, ovarian cancer transplanted i.m. on BDF1 mice and treatment byintraperitoneal route. Dose: % mg/kg Tumour Tumour Tumour Tumour TumourTumour Mean Tumour body weight weight weight weight weight weight tumourgrowth Days of weight (g) (g) (g) (g) (g) (g) weight sem inhibitiontreatment 400 0 0 0 0 0 0.052 0.008 0.008 97 1-11 Control 0.591 0.2350.032 0.162 0.449 0.288 0.292 0.082 0 untreated

TABLE 3 The effect of the compound of Example 3 on the growth of MAC15tumour, murine colon cancer, transplanted s.c. on NMRI mice andtreatment given by intraperitoneal route. Dose: % mg/kg Tumour TumourTumour Tumour Tumour Tumour Mean Tumour body weight weight weight weightweight weight tumour growth Days of weight (g) (g) (g) (g) (g) (g)weight sem inhibition treatment 400 0.074 0.094 0.190 0.096 0.132 —0.117 0.02 81 1-5 300 0.194 0.201 0.190 0.176 0.214 — 0.195 0.006 68 1-5Control 0.581 0.617 0.770 0.612 0.417 — 0.610 0.056 0 — untreated

TABLE 4 The effect of the compound of Example 4 on the growth of MAC15tumour, murine colon cancer, transplanted s.c. on NMRI mice andtreatment given by intraperitoneal route from day 1-5. Dose: % mg/kgTumour Tumour Tumour Tumour Tumour Tumour Tumour Tumour Tumour TumourMean Tumour body weight weight weight weight weight weight weight weightweight weight tumour growth weight (g) (g) (g) (g) (g) (g) (g) (g) (g)(g) weight sem inhibition 500 0.076 0.033 0.0 0.090 0.060 0.0 0.1340.114 0.0 — 0.056 0.017 86 Control 0.342 0.518 0.437 0.312 0.444 0.3280.561 0.239 0.340 0.459 0.398 0.032 0 untreated

EXAMPLE 6

Antitumour Test Protocol and Results for H460 Non-Small Cell Lung(NSCLC) and DLD-1 colon cancers human Xenograft.

Fragments of the non-small cell lung cancer, H460, and DLD-1 coloncancer human xenografts were transplanted sub-cutaneously in both flanksof 14 female Nu nude mice from an inbred colony (B7K Universal, Hull,UK). Mice received CRM diet (SDS, WItham, UK) and water ad libitum. Allexperiments were carried out under a project license issued by the UKHome Office, and UKCCCR guidelines were followed throughout.

Therapy commenced once tumours had grown to a sufficient size to bemeasured accurately (minimum 4 mm×4 mm), which was 10 days for H460 and19 days for DLD-1. 10 tumours grew sufficiently to use in 7 mice forboth the test and untreated groups.

In the case of H460, the compound of Example 4 was administered at aconcentration of 500 mg/kg body weight/day, intraperitoneally (i.p.),daily for 5 days (designated days 0-4). Animals were monitored daily forany toxicity effects, and tumours were measured, and weights recordeddaily over the period of therapy and then on day 7 (untreated and testgroups), and day 8 (test group). On day 7 for the untreated group, andday 8 for the test group, the largest tumour diameter was greater thanthe maximum permitted 17 mm, and subjects were then euthenised by UKHome Office Schedule 1 methods.

Antitumour activity against DLD-1 was assessed by dosing i.p. daily for5 days (500mg/kg body weight/day) followed by a 2 day break, and thendosing for a further 5 days, and assessing tumour growth compared to acontrol untreated group until the tumours have reached the maximumpermitted size (day 16 control and tested group).

In both cases tumour volumes were assessed by caliper measurements of 2perpendicular diameters and the volumes estimated using the followingequation:Volumes−(a²×b)/2,where a is the smaller, and b the larger diameter of the two. Theeffects of the therapy were determined from the difference in tumourvolume-doubling time between treated and control groups, and thesignificance of any tumour growth delay determined using theMann-Whitney U Test.

The results are set out in Tables 5 and 6 below.

1. H460 NSCLC xenograft TABLE 5 Mean time Median time Growth MaximumGroup to RTV2 RTV2 delay % weight Number (days) (days) (days)Significance loss 1 2.6 2.5 — — 4.9 2 6.2 6.4 3.9 P < 0.01 6.9GROUP 1 - untreated controlsGROUP 2 - Compound of Example 4, 500 mg/kg, i.p. days 0-4

2. DLD-1 colon cancer xenograft. TABLE 6 Mean time Median time GrowthMaximum Group to RTV2 RTV2 delay % weight Number (days) (days) (days)Significance loss 1 3.2 3.1 — — 3.2 2 9.9 9.8 6.7 P < 0.01 4.3GROUP 1 - untreated controlsGROUP 2 - Compound of Example 4, 500 mg/kg, i.p. days 0-4RTV2 = Relative Tumour Volume Doubling.

1-7. (canceled)
 8. A compound of the general formula IIA or IIB

or a salt thereof, in which R¹ represents a hydrogen atom or anoptionally substituted alkyl, alkenyl, alkynyl, cycloalkyl,cycloalkenyl, aryl, aralkyl, heterocyclyl or heterocyclylalkyl group; R²represents a hydrogen atom or an optionally substituted alkyl, alkenyl,alkynyl, cycloalkyl, cycloalkenyl, aryl, aralkyl, acyl, heterocyclyl orheterocyclylalkyl group; and W and X each independently represents ahydrogen or halogen atom and Y represents a hydrogen atom, a hydroxylgroup or a group —OM where M represents an alkali metal atom,
 9. Acompound according to claim 8 in which Y represents a hydrogen atom, ahydroxyl group or a group —OM where M represents a sodium or potassiumatom.
 10. A compound according to claim 8 in which R¹ represents amethyl group, R² represents a hydrogen atom, W and X both represent ahydrogen atom or both represent a chlorine atom, and Y represents ahydroxyl group. 11.(Z)-2,3-dichloro-4-[N-formyl-N-methylamino]-4-oxo-but-2-enoic acid. 12.A process for the preparation of a compound according to claim 8, whichcomprises reacting with a base a compound of the general formula

in which R¹, R², W and X are as defined in claim 8 and R³ represents anoptionally substituted alkyl, aryl or aralkyl group. 13-17. (canceled)18. A pharmaceutical composition comprising a carrier and one or morecompounds according to claim
 8. 19. A method for treating proliferativediseases, microbial diseases or viral diseases comprising administeringto a patient a therapeutically effective amount of a compound accordingto claim
 8. 20. A method according to claim 19 for inhibiting tumourgrowth.
 21. A compound according to claim 8 in which R¹ represents aC₁₋₁₂ alkyl or C₆₋₁₄ aryl group, each group being optionally substitutedby one or more substituents selected from the group consisting ofhalogen atoms, nitro, cyano, hydroxyl, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄alkoxy and C₁₋₄ haloalkoxy groups.
 22. A compound according to claim 8in which R² represents a hydrogen atom or a C1-12 alkyl or C6-14 arylgroup, each group being optionally substituted by one or moresubstituents selected from the group consisting of halogen atoms, nitro,cyano, hydroxyl, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxy and C₁₋₄haloalkoxy groups.
 23. A compound according to claim 8 in which W and Xeach independently represents a hydrogen, chlorine or bromine atom.